The overall goal of this application is to develop improved plague vaccines. Our proposal was developed based on our recent progress in plague vaccine studies which includes 1.) the use of a DNA immunization approach to search for additional protective antigens and 2.) the discovery of cell mediated immune responses in the protection of Y. pestis infection. Support for the current proposal will allow us to develop a unique technological platform which can combine antigen discovery, optimization of vaccine formulation, and efficient needle free intradermal delivery of subunit-based multi-gene plague vaccines into one system. Specific Aim 1: To continue our work using DNA immunization as a tool to screen for additional protective antigens against plague, including both the study of individual candidate antigens and the high throughput screening of animal sera which has been immunized with specialized Y. pestis strains deficient for the dominant antigens LcrV and F1. This approach exploits the availability of a newly constructed Y. pestis expression library and protein microarray techniques. Specific Aim 2: To study the cellular immunological mechanisms that are important for improving the protective efficacy of plague vaccines which includes the use of the DNA prime plus protein boost approach to increase the level and longevity of protective antibody responses induced by subunit plague vaccines. Specific Aim 3: To conduct late-phase preclinical immunogenicity and protection studies in non-human primates to prepare for a clinical study with optimized plague vaccine formulation and DNA delivery device that has been proven effective in humans. Plain, lay language summary: This study is designed to produce effective biodefense vaccines against Y. pestis, the highly lethal bacteria that causes plague, an infection that has the potential to cause a high percentage of human fatalities. This study is part of the broad biodefense effort to protect against such agents if they were to be used for bioterrorism purposes.